Methods of treating a subject having heart failure

ABSTRACT

Methods of treating a subject having a cardiovascular disease, selecting a therapy for a subject having a cardiovascular disease, identifying a subject having a cardiovascular disease that will benefit or not benefit from exercise therapy, determining whether a subject having a cardiovascular disease should begin, continue, not begin, discontinue, or avoid exercise therapy, determining whether a subject having a cardiovascular disease should continue, discontinue, or avoid exercise therapy, reducing the risk of an adverse outcome (e.g., death) in a subject having a cardiovascular disease, and predicting the efficacy of exercise therapy in a subject having a cardiovascular disease. These methods include determining a level of soluble ST2 in a subject.

CLAIM OF PRIORITY

This application is a continuation of U.S. patent application Ser. No.13/552,553, filed on Jul. 18, 2012 (issued as U.S. Pat. No. 8,748,110),which claims the benefit of U.S. Provisional Patent Application Ser. No.61/508,923, filed on Jul. 18, 2011, and U.S. Provisional PatentApplication Ser. No. 61/509,359, filed on Jul. 19, 2011, the entirecontents of which are herein incorporated by reference.

TECHNICAL FIELD

This invention relates to methods of treating cardiovascular diseasesusing exercise therapy, and predicting the efficacy of exercise therapy.

BACKGROUND

Circulating biomarkers play a critical role in the diagnosis andmanagement of patients with chronic heart failure (Braunwald, N. Engl.J. Med. 358:2148-2159, 2008). Natriuretic peptides, such as brainnatriuretic peptide (BNP) and N-terminal pro-BNP (NT-proBNP), have beendemonstrated to be powerful tools for the diagnosis, riskstratification, and management of patients with heart failure (Felker etal., Canadian Med. Assoc. J. 175:611-617, 2006). In addition to beinguseful for clinical management, biomarkers can provide insights into themechanisms underlying important physiologic relationships. Exerciseintolerance, typically manifested as exertional dyspnea, is a majormorbidity of chronic heart failure. Both maximal (e.g., as measured bypeak oxygen uptake [peak VO₂]) (Aaronson et al., Circulation95:2660-2667, 1997; Mancini et al., Circulation 83:778-786, 1991) andsubmaximal exercise capacity (e.g., as measured by distance in the6-minute walk test) (Bittner et al., JAMA 270:1702-1707, 1993) have beendemonstrated to be of substantial prognostic importance in chronic heartfailure.

A variety of therapies can be used to treat patients diagnosed with acardiovascular disease. For example, exercise therapy is commonly usedto treat patients diagnosed with a cardiovascular disease (see, forexample, Korhonen et al., J. Womens Health 20:1051-1064, 2011).

SUMMARY

Applicants have discovered a correlation between the level of solubleST2 and the efficacy of exercise therapy in subjects having acardiovascular disease, and a correlation between soluble ST2 level,exercise therapy, and the risk of an adverse outcome (e.g., death) in asubject having a cardiovascular disease.

Provided herein are methods of treating a subject having acardiovascular disease that, in some embodiments, include determining alevel of soluble ST2 in a biological sample from the subject,identifying a subject that has a decreased level of soluble ST2 in thebiological sample compared to a risk reference level of soluble ST2, andselecting the identified subject for exercise therapy. Also provided aremethods of treating a subject having a cardiovascular disease thatinclude determining a level of soluble ST2 in a biological sample fromthe subject, identifying a subject that has an elevated level of solubleST2 in the biological sample compared to a risk reference level ofsoluble ST2; and instructing the identified subject not to begin, todiscontinue, or to avoid exercise therapy.

Also provided are methods of selecting a therapy for a subject having acardiovascular disease that include determining a level of soluble ST2in a biological sample from the subject, and comparing the level ofsoluble ST2 in the biological sample to a risk reference level ofsoluble ST2, where a decreased level of soluble ST2 in the biologicalsample compared to the risk reference level indicates that the subjectshould begin or continue exercise therapy, and an elevated level ofsoluble ST2 in the biological sample compared to the risk referencelevel indicates that the subject should not begin or should discontinueexercise therapy.

Also provided are methods of identifying a subject having acardiovascular disease that will benefit from exercise therapy thatinclude determining a level of soluble ST2 in a biological sample fromthe subject, and selecting a subject that has a decreased level ofsoluble ST2 in the biological sample as compared to a risk referencelevel of soluble ST2, where the selected subject is identified as asubject that will benefit from exercise therapy. Also provided aremethods of identifying a subject having a cardiovascular disease thatwill not benefit from exercise therapy that include determining a levelof soluble ST2 in a biological sample from the subject, and selecting asubject that has an elevated level of soluble ST2 in the biologicalsample compared to a risk reference level of soluble ST2, where theselected subject is identified as a subject that will not benefit fromexercise therapy.

Also provided are methods of determining whether a subject having acardiovascular disease should begin, continue, not begin, or discontinueexercise therapy that include determining a level of soluble ST2 in abiological sample from the subject, where a decreased level of solubleST2 in the biological sample compared to a risk reference level ofsoluble ST2 indicates that the subject should begin or continue exercisetherapy, and an elevated level of soluble ST2 indicates that the subjectshould not begin or discontinue exercise therapy.

Also provided are methods of determining whether a subject having acardiovascular disease should discontinue or continue exercise therapythat include determining a level of soluble ST2 in a biological samplefrom the subject at a first time point before or after the start ofexercise therapy, and determining a level of soluble ST2 in a biologicalsample from the subject undergoing exercise therapy at a second timepoint after the start of exercise therapy and after the first timepoint, where an elevation in the level of soluble ST2 in the biologicalsample at the second time point compared to the level of soluble ST2 inthe biological sample at the first time point indicates that the subjectshould discontinue exercise therapy, and a decrease in the level ofsoluble ST2 in the biological sample at the second time point comparedto the level of soluble ST2 in the biological sample at the first timepoint indicates that the subject should continue exercise therapy.

In some embodiments of the methods described herein, the methods includedetermining that the subject has a level of soluble ST2 that is above afirst reference level (e.g., a first level indicating that the subjecthas a cardiovascular disease, or is at risk of an adverse cardiovascularevent; e.g., as described in U.S. Pat. No. 7,998,683; US2011/0262941;US2012/0040381; U.S. Pat. No. 8,090,562; US2012/0065897; U.S. Pat. Nos.7,670,769; 7,655,415; 7,989,210; US2011/0250703; U.S. Pat. Nos.7,432,060; 7,985,558; US2011/028088, all of which are incorporatedherein by reference) and below a second, risk reference level (e.g.,below a level indicating that the subject is at risk of an adverse eventif they engage in exercise, as described herein), and the subject isselected for exercise therapy. Thus in some embodiments, the methodsinclude determining that the subject has a level of ST2 that fallswithin a range that is associated with the presence of a cardiovasculardisease that would benefit from exercise therapy and the absence of highrisk of an adverse event associated with exercise therapy.

Also provided are methods of reducing the risk of an adverse outcome ina subject having a cardiovascular disease that include determining alevel of soluble ST2 in a biological sample from the subject,identifying a subject that has a decreased level of soluble ST2 in thebiological sample compared to a risk reference level of soluble ST2, andselecting the identified subject for exercise therapy. Also provided aremethods of reducing the risk of an adverse outcome in a subject having acardiovascular disease that include determining a level of soluble ST2in a biological sample from the subject, identifying a subject that hasan elevated level of soluble ST2 in the biological sample compared to arisk reference level of soluble ST2, and instructing the subject to notbegin or to discontinue exercise therapy. In some embodiments of thesemethods, the risk of adverse outcome is risk of death.

Also provided are methods of predicting the efficacy of exercise therapyin a subject having a cardiovascular disease that include determining alevel of soluble ST2 in a biological sample from the subject, andcomparing the level of soluble ST2 in the biological sample to anefficacy reference level of soluble ST2, where a decreased level ofsoluble ST2 in the biological sample compared to the efficacy referencelevel of soluble ST2 indicates that the exercise therapy will beeffective in the subject, and an elevated level of soluble ST2 in thebiological sample compared to the efficacy reference level of solubleST2 indicates that the exercise therapy will not be effective in thesubject.

In any of the methods described herein, the biological sample containsblood or serum. In any of the methods described herein, the determiningis performed using an antibody or an antibody fragment that binds tosoluble ST2. In any of the methods described herein, the reference levelof soluble ST2 is a predetermined threshold value. In any of the methodsdescribed herein, the reference level of soluble ST2 is a level ofsoluble ST2 in a healthy subject. In any of the methods describedherein, the subject is hyperchlosterolemic, hypertriglyceridemic,hyperlipidemic, a smoker, hypertensive, or has a body mass index ofgreater than 30. Some embodiments of the methods described hereinfurther include determining a level of cardiac troponin I, B-typenatriuretic peptide, atrial natriuretic peptide, or C-reactive proteinin the biological sample. Some embodiments of the methods describedherein further include determining a level of level of cardiac troponinI, B-type natriuretic peptide, atrial natriuretic peptide, or C-reactiveprotein in the biological sample at the first time point or thebiological sample at the second time point.

In some embodiments of any of the methods described herein, thecardiovascular disease is selected from the group of: cardiachypertrophy, myocardial infarction, stroke, arteriosclerosis, and heartfailure. In some embodiments of any of the methods described herein, thesubject is administered at least one therapeutic agent selected from thegroup of: anti-inflammatory agents, anti-thrombotic agents,anti-coagulants, anti-platelet agents, lipid-reducing agents (e.g., astatin), direct thrombin inhibitors, glycoprotein IIb/IIIb receptorinhibitors, calcium channel blockers, beta-adrenergic receptor blockers,cyclooxygenase-2 inhibitors, and renin-angiotensin-aldosterone system(RAAS) inhibitors. In some embodiments, the RAAS inhibitor is selectedfrom the group of: an angiotensin-converting enzyme inhibitor, anangiotensin II receptor blocker, and an aldosterone antagonist.

As used herein, the term “cardiovascular disease” refers to a disorderof the heart and blood vessels, and includes disorders of the arteries,veins, arterioles, venules, and capillaries. Non-limiting examples ofcardiovascular diseases include cardiac hypertrophy, myocardialinfarction, stroke, arteriosclerosis, and heart failure. Additionalexamples of cardiovascular diseases are known in the art.

By the term “soluble ST2” is meant a soluble protein containing asequence at least 90% identical (e.g., at least 95%, 96%, 97%, 98%, 99%,or 100% identical) to NCBI Accession No. NP_003847.2 (SEQ ID NO: 1) orcontaining a sequence at least 90% identical (e.g., at least 95%, 96%,97%, 98%, 99%, or 100% identical) to amino acids 19-328 of SEQ ID NO: 1,or a nucleic acid containing a sequence at least 90% identical (e.g., atleast 95%, 96%, 97%, 98%, 99%, or 100% identical) to NCBI Accession No.NM_003856.2 (SEQ ID NO: 2) or containing a sequence at least 90%identical (e.g., at least 95%, 96%, 97%, 98%, 99%, or 100% identical) tonucleotides 285 to 1214 of SEQ ID NO: 2.

By the term “elevated” or “elevation” is meant a difference, e.g., thepresence of a statistically significant or detectable increase in adetermined or measured level (e.g., a human soluble ST2 protein level)compared to a reference level (e.g., a level of human soluble ST2 in asubject not having a disease, a subject not presenting with two or moresymptoms of a disease, or a subject not identified as being at risk ofdeveloping a disease, or a threshold level of human soluble ST2). Insome embodiments, the reference is a threshold level, and any levelabove that is considered “elevated.” Additional reference levels ofhuman soluble ST2 are described herein and are known in the art.

By the term “reference level” is meant a threshold level or a level in acontrol subject or control patient population. A reference level willdepend on the assay performed and can be determined by one of ordinaryskill in the art. Non-limiting examples of reference levels aredescribed herein and are known in the art. Reference levels of humansoluble ST2 can be determined using methods known in the art.

In some embodiments, the reference level is a risk reference level,e.g., a risk reference level of soluble ST2 in a subject who experiencedor was more likely to experience an adverse outcome and engaged inexercise, a level in a population of subjects who experienced or weremore likely to experience an adverse outcome and engaged in exercise, ora threshold level of soluble ST2 above which the risk of an adverseoutcome is increased in those who engange in exercise therapy.

In some embodiments, the reference level is an efficacy reference level,e.g., an efficacy reference level of soluble ST2 is a level in a subjectwho experienced a therapeutic benefit from exercise therapy, a level ina population of subjects who experienced a therapeutic benefit fromexercise therapy, or a threshold level of soluble ST2 below which thesubject is likely to experience a therapeutic benefit from exercisetherapy.

By the term “additional marker” is meant a protein, nucleic acid, lipid,or carbohydrate, or a combination (e.g., two or more) thereof, that isdiagnostic of the presence of a particular disease. The methodsdescribed herein can include detecting the level of soluble human ST2and at least one additional marker in a biological sample from asubject. Non-limiting examples of additional markers that can bedetected include: proANP, NT-proANP, ANP, proBNP, NT-proBNP, BNP,troponin, CRP, creatinine, Blood Urea Nitrogen (BUN), liver functionenzymes, albumin, and bacterial endotoxin; and those markers describedin U.S. Patent Application Publication Nos.: 2007/0248981; 2011/0053170;2010/0009356; 2010/0055683; and 2009/0264779 (each of which is herebyincorporated by reference).

By the term “hypertriglyceridemia” is meant a triglyceride level that isgreater than or equal to 180 ng/mL (e.g., greater than or equal to 200ng/mL).

By the term “hypercholesterolemia” is meant an increased level of atleast one form of cholesterol or total cholesterol in a subject. Forexample, a subject with hypercholesterolemia can have a high densitylipoprotein (HDL) level of ≧40 mg/dL (e.g., >50 mg/dL or >60 mg/mL), alow density lipoprotein (LDL) level of >130 mg/dL (e.g., >160 mg/dLor >200 mg/dL), and/or a total cholesterol level of >200 mg/dL (e.g.,240 mg/dL).

By the term “hypertension” is meant an increased level of systolicand/or diastolic blood pressure. For example, a subject withhypertension can have a systolic blood pressure that is >120 mmHg(e.g., >140 mmHg or >160 mmHg) and/or a diastolic blood pressure thatis >80 mmHg (e.g., >90 mmHg or >100 mmHg).

By the term “healthy subject” is meant a subject that does not have adisease (e.g., cardiovascular disease). For example, a healthy subjecthas not been diagnosed as having a disease and is not presenting withtwo or more (e.g., two, three, four, or five) symptoms of a diseasestate.

By “risk of death” is meant the risk of death in a subject from adisease or complications associated with a disease (e.g., acardiovascular disease) compared to a reference population. The termrisk of death as used herein excludes intentional or accidental death,e.g., death by blunt or crushing trauma, such as a car accident.

As used herein, a “biological sample” includes one or more of blood,serum, plasma, urine, and body tissue. Generally, a biological sample isa sample containing serum, blood, or plasma.

By the term “statin” is meant a therapeutic molecule that inhibits theenzyme HMG-CoA reductase. Non-limiting examples of statins include:atorvastatin, fluvastatin, lovastatin, pitavastatin, pravastatin,rosuvastatin, and simvastatin. Additional examples of statins are knownin the art.

By “adverse outcome” is meant any detrimental event that occurs in asubject as a result of a disease (e.g., a cardiovascular disease).Non-limiting examples of adverse outcomes in a subject having acardiovascular disease include: organ failure, organ transplantation,hospitalization or rehospitalization, recurrence of one or more symptomsof a cardiovascular disease, development of one or more additionalsymptoms of a cardiovascular disease, an increase in the frequency,intensity, or duration of one or more symptoms of a cardiovasculardisease experienced by the subject, a first or subsequent myocardialinfarction, or death (mortality). In preferred embodiments, the adverseoutcome is mortality.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Methods and materials aredescribed herein for use in the present invention; other, suitablemethods and materials known in the art can also be used. The materials,methods, and examples are illustrative only and not intended to belimiting. All publications, patent applications, patents, sequences,database entries, and other references mentioned herein are incorporatedby reference in their entirety. In case of conflict, the presentspecification, including definitions, will control.

The details of one or more embodiments of the invention are set forth inthe accompanying drawings and the description below. Other features,objects, and advantages of the invention will be apparent from thedescription and drawings, and from the claims.

DESCRIPTION OF DRAWINGS

FIG. 1 is a Kaplan-Meier graph showing the surviving proportion of heartfailure subjects over time within the identified groups. The survivaldata for subjects having a soluble ST2 level less than or equal to 20.4ng/mL (top line); greater than 20.4 ng/mL, but less than or equal to28.6 ng/mL (middle line); and greater than 28.6 ng/mL (bottom line) areshown (log-rank test, p<0.001).

FIG. 2 is a graph and table showing the percentage of mortality in heartfailure subjects over time. The percentage mortality data for heartfailure subjects having a soluble ST2 level equal to or less than 35ng/mL (left bar for each time point), or greater than 35 ng/mL (rightbar for each time point) are shown. The table also shows the relativerisk of mortality at each time point for subjects having a soluble ST2level greater than 35 ng/mL compared to subjects having a soluble ST2level less than or equal to 35 ng/mL.

FIG. 3 is a Kaplan-Meier graph showing the surviving proportion of heartfailure subjects over time within the identified groups. The survivaldata for subjects having a soluble ST2 level equal to or less than 35ng/mL and performing an exercise treatment regimen (top line), subjectshaving a level of soluble ST2 greater than 35 ng/mL and not performingan exercise treatment regimen (second line from top), subject having alevel of soluble ST2 greater than 35 ng/mL and not performing anexercise treatment regime (second line from bottom), and subjects havinga level of soluble ST2 equal to or less than 35 ng/mL and performing anexercise treatment regime (bottom line) are shown.

FIG. 4 is a graph of the hazard ratio for mortality within 1 year inheart failure subjects performing an exercise treatment regime comparedto heart failure subjects not performing an exercise treatment regimehaving the different soluble ST2 levels shown.

FIG. 5 shows the protein sequence of Soluble Human ST2. Signal PeptideAmino acids 1-18 (underlined); Mature Peptide after removal of signalsequence amino acids (19-328).

FIG. 6 shows the mRNA sequence of soluble human ST2. Nucleotides 285 to1214 encode the amino acid sequence (without the signal sequence) ofsoluble human ST2.

FIGS. 7A-B shows the protein sequence of the long form of human St2(membrane-bound). Signal Peptide Amino acids 1-18 (underlined); MaturePeptide after removal of signal sequence amino acids (19-556).

FIG. 8 shows the mRNA sequence of human ST2. Nucleotides 326 to 1939encode the amino acid sequence (without the signal sequence) of the longform of human ST2.

DETAILED DESCRIPTION

Provided herein are methods of treating a subject having acardiovascular disease; selecting a therapy for a subject having acardiovascular disease; identifying a subject that will benefit or willnot benefit from exercise therapy; determining whether a subject havinga cardiovascular disease should begin, continue, or discontinue exercisetherapy; and predicting the efficacy of exercise therapy in a subjecthaving a cardiovascular disease. Also provided are methods of reducingthe risk an adverse outcome (e.g., death) in a subject having acardiovascular disease. These methods require determining a level ofsoluble ST2 in a biological sample from the subject.

Cardiovascular Diseases

A cardiovascular disease is a disorder of the heart and blood vessels(e.g., disorders of the arteries, veins, arterioles, venules, andcapillaries). Cardiovascular diseases can be diagnosed using methodsknown in the art. Non-limiting examples of cardiovascular diseaseinclude congestive heart failure, stroke, acute coronary artery disease,arrhythmia, asymmetric septal hypertrophy (e.g., left ventricularhypertrophy with resultant diastolic dysfunction), cardiomyopathy,valvular dysfunction, pericarditis, atherosclerosis, and myocardialinfarction. A subject can be diagnosed as having a cardiovasculardisease by a medical professional (e.g., a physician, a physician'sassistant, a nurse, a nurse's assistant, or a laboratory technician)using exemplary methods described herein. Additional methods fordiagnosing a cardiovascular disease are known in the art.

Heart failure is a clinical syndrome of diverse etiologies linked by thecommon feature of impaired heart pumping and characterized by thefailure of the heart to pump blood commensurate with the requirements ofthe metabolizing tissues. Heart failure can be diagnosed in a subject bythe observation of one or more of the following non-limiting symptoms ina subject: dyspnea, fatigue and weakness, edema in the legs, ankles, andfeet, rapid or irregular heartbeat, reduced ability to exercise,persistent cough or wheezing, white or pink blood-tinged phlegm,abdominal swelling (ascites), sudden weight gain from fluid retention,lack of appetite, nausea, difficulty concentrating or decreasedalertness, and chest pain. Additional non-limiting methods fordiagnosing heart failure in a subject include the use of commerciallyavailable diagnostic tests (e.g., enzyme-linked immunosorbent assays)known in the art.

A myocardial infarction is a focus of necrosis resulting from inadequateperfusion of the cardiac tissue. Myocardial infarction generally occursfrom an abrupt decrease in coronary blood flow that follows a thromboticocclusion of a coronary artery previously narrowed by atherosclerosis.Generally, a myocardial infarction occurs when an atherosclerotic plaquefissures, ruptures, or ulcerates, and a mural thrombus forms leading tocoronary artery occlusion. Non-limiting methods of diagnosing amyocardial infarction include the use of a number of commerciallyavailable diagnostic tests known in the art. Generally, these diagnostictests may be divided into four main categories: (1) nonspecific indexesof tissue necrosis and inflammation; (2) electrocardiograms; (3) serumenzyme changes (e.g., creatine phosphokinase levels); and (4) cardiacimaging. A myocardial infarction can also be diagnosed by theobservation of one or more of the following symptoms in a subject: chestpain (typically on the left side of the body), neck or jaw pain,shoulder or arm pain, clammy skin, dyspnea, nausea, and vomiting.Additional methods of diagnosing a myocardial infarction are known inthe art.

A stroke can be diagnosed in a subject by the observation of one or moresymptoms and/or by a physical examination (e.g., interventional andnon-interventional diagnostic tools, such as computed tomography andmagnetic resonance imaging). Non-limiting symptoms of a stroke include:paralysis, weakness, decreased sensation and/or vision, numbness,tingling, aphasia (e.g., inability to speak or slurred speech, ordifficulty reading or writing), agnosia (i.e., inability to recognize oridentify sensory stimuli), loss of memory, coordination difficulties,lethargy, sleepiness or unconsciousness, lack of bladder or bowelcontrol, and cognitive decline (e.g., dementia, limited attention span,and inability to concentrate). In some examples, medical imagingtechniques can be used identify a subject having an infarct or ahemorrhage in the brain.

Cardiac hypertrophy is typically characterized by left ventricularhypertrophy, usually of a non-dilated chamber, that occurs without anobvious antecedent cause. Cardiac hypertrophy can be diagnosed throughthe use of electrocardiography or echocardiography.

Arteriosclerosis is a cardiovascular disease characterized by ahardening or loss of elasticity. Arteriosclerosis can be diagnosed bythe detection of one or more of the following physical symptoms: a weakor absent pulse below the narrowed area of an artery, decreased bloodpressure in an affected limb, bruits caused by turbulent flow in anartery, an aneurysm in the abdomen or behind the knee, poor woundhealing, and increased levels of cholesterol. Atherosclerosis can alsobe detected using imaging techniques including, but not limited to:Doppler ultrasound, electrocardiography, angiography, computedtomography, or magnetic reasonance (e.g., magnetic reasonanceangiography).

A subject can be diagnosed as having a cardiovascular disease followingadmission to a hospital or following presentation to a health careclinic. Effective therapeutic treatment of a cardiovascular disease canbe determined by observing a decrease in the number of symptoms of acardiovascular disease in a subject or a decrease in the frequency,intensity, and/or duration of one or more symptoms of a cardiovasculardisease (e.g., any of the symptoms described herein) in a subject.Effective therapeutic treatment of a cardiovascular disease can also bedetermined by detecting a decrease in the levels of one or more markersof a cardiovascular disease (e.g., any of the markers of cardiovasculardisease known in the art or described herein) in a biological samplefrom a subject over time (e.g., a significant decrease in the level ofat least one marker at a second time point compared to the level of thebiomarker at a first time point prior to the start of treatment or at anearlier time point during the treatment period). Successful treatment ofa cardiovascular disease can also be determined by a decreased risk ofan adverse event (e.g., a decreased risk of death, hospitalization orrehospitalization, organ failure, organ transplantation, or a first orsubsequent myocardial infarction) (e.g., compared to the risk of anadverse event in patient population diagnosed with the samecardiovascular disease but receiving no treatment or a differenttreatment).

Exercise Therapy

Subjects diagnosed as having a cardiovascular disease are oftendirected/instructed by a healthcare provider to perform an exercisetherapy regime. Exercise therapy is a treatment regime that involves theperiodic performance of physical activity by a subject (e.g., a subjectdiagnosed as having a cardiovascular disease). The physical activityperformed during this type of therapy can be aerobic (e.g., walking,jogging/running, swimming, biking, or rowing) or anaerobic (e.g., weightlifting or resistance training) exercise. The physical activity can beperformed in the presence of a health care professional (e.g., aphysical therapist, a nurse, a nurse's assistant, a physician'sassistant, or a physician). The physical activity can be performed atleast once a week (e.g., at least once a day, twice a day, two times aweek, three times a week, four times a week, five times a week, or sixtimes a week). In some embodiments, a single episode of physicalactivity within the exercise therapy regime can last between 5 minutesand 3 hours, between 5 minutes and 2 hours, between 5 minutes and 1hour, or between 10 minutes and 1 hour. The intensity and/or type of thephysical activity performed by the subject can vary with the physicalcondition of the subject (e.g., age, severity of cardiovascular disease,additional disease states, weight, and blood pressure). A subject cancontinue to perform an exercise therapy regime over an extended periodof time (e.g., over a period of 1 month to 1 year, 1 month to 2 years, 1year to 3 years, 2 years to 5 years, or 4 years to 10 years). In someembodiments, a subject can be monitored by a health care professional toadjust one or more parameters of the exercise therapy regime, includingthe frequency, intensity, length of individual episodes of physicalactivity, and the type of physical activity performed. The efficacy ofexercise therapy in a subject having a cardiovascular disease can bedetermined using any of the methods for determining successful treatmentof a cardiovascular disease (e.g., those methods described herein orknown in the art). A subject can continue to perform an exercise therapyregime until instructed by a health care professional to discontinue theexercise therapy or until a specific therapeutic outcome has beenachieved (e.g., a decrease in the number of symptoms of a cardiovasculardisease has been achieved, a decrease in severity, intensity, orfrequency of one or more symptoms of a cardiovascular disease has beenachieved, or a decrease in the levels of one or more markers of acardiovascular disease has been achieved in the subject).

ST2

The ST2 gene is a member of the interleukin-1 receptor family whoseprotein product exists both as a trans-membrane form as well as asoluble receptor that is detectable in serum (Kieser et al., FEBS Lett.372(2-3):189-193, 1995; Kumar et al., J. Biol. Chem.270(46):27905-27913, 1995; Yanagisawa et al., FEBS Lett. 302(1):51-53,1992; Kuroiwa et al., Hybridoma 19(2):151-159, 2000). Soluble ST2 wasdescribed to be markedly up-regulated in an experimental model of heartfailure (Weinberg et al., Circulation 106(23):2961-2966, 2002), and datasuggest that human soluble ST2 concentrations are also elevated in thosewith chronic severe heart failure (Weinberg et al., Circulation107(5):721-726, 2003), as well as in those with acute myocardialinfarction (Shimpo et al., Circulation 109(18):2186-2190, 2004).

Without wishing to be bound by theory, the transmembrane form of ST2 isthought to play a role in modulating responses of T helper type 2 cells(Lohning et al., Proc. Natl. Acad. Sci. U.S.A. 95(12):6930-6935, 1998;Schmitz et al., Immunity 23(5):479-490, 2005), and may play a role indevelopment of tolerance in states of severe or chronic inflammation(Brint et al., Nat. Immunol. 5(4):373-379, 2004), while the soluble formof ST2 is up-regulated in growth stimulated fibroblasts (Yanagisawa etal., 1992, supra). Experimental data suggest that the ST2 gene ismarkedly up-regulated in states of cardiomyocyte stretch (Weinberg etal., 2002, supra) in a manner analogous to the induction of the BNP gene(Bruneau et al., Cardiovasc. Res. 28(10):1519-1525, 1994).

Tominaga et al. (FEBS Lett. 258:301-304, 1989) isolated murine genesthat were specifically expressed by growth stimulation in BALB/c-3T3cells. Haga et al. (Eur. J. Biochem. 270:163-170, 2003) describes thatthe ST2 gene was named on the basis of its induction by growthstimulation. The ST2 gene encodes two protein products: ST2 or sST2,which is a soluble secreted form, and ST2L, a transmembrane receptorform that is very similar to the interleukin-1 receptors. The HUGONomenclature Committee designated the human homolog of ST2, the cloningof which was described in Tominaga et al., Biochim. Biophys. Acta.1171:215-218, 1992, as Interleukin 1 Receptor-Like 1 (IL1RL1). The twoterms are used interchangeably in the art.

The mRNA sequence of the shorter, soluble isoform of human ST2 can befound at GenBank Acc. No. NM_003856.2 (SEQ ID NO: 2), and thepolypeptide sequence is at GenBank Acc. No. NP_003847.2 (SEQ ID NO: 1).The mRNA sequence for the longer form of human ST2 is at GenBank Acc.No. NM_016232.4 (SEQ ID NO: 4), and the polypeptide sequence is atGenBank Acc. No. NP_057316.3 (SEQ ID NO: 3). Additional information isavailable in the public databases at GeneID: 9173, MIM ID #601203, andUniGene No. Hs.66.

Methods for detecting and measuring soluble ST2 are known in the art,e.g., as described in U.S. Patent Application Publication Nos.2003/0124624, 2004/0048286, and 2005/0130136, and U.S. patentapplication Ser. No. 13/083,333 and PCT Application No.PCT/US2011/031801, the entire contents of which are incorporated hereinby reference. These U.S. patent application publications describemethods of determining the level of soluble ST2 using an antibody orantibody fragment that binds to soluble ST2.

In some embodiments, the antibody is a monoclonal antibody produced bythe hybridoma deposited at the ATCC and designated by Patent DepositDesignation PTA-10431 (the 7E4 antibody), or is an antigen-bindingfragment of the antibody produced by the hybridoma deposited at the ATCCand designated by the Patent Deposit Designation PTA-10431 (fragments ofthe 7E4 antibody). In some embodiments, the antibody is a monoclonalantibody produced by the hybridoma deposited at the ATCC and designatedby Patent Deposit Designation PTA-10432 (the 9F8 antibody), or is anantigen-binding fragment of the antibody produced by the hybridomadeposited at the ATCC and designated by the Patent Deposit DesignationPTA-10432 (fragments of the 9F8 antibody). Combinations of two or moreof the antibodies or fragments described herein (e.g., two or more of a7E4 antibody, 7E4 antibody fragments, 9F8 antibody, and 9F8 antibodyfragments) are useful in any of the methods described herein.

The human soluble ST2-binding monoclonal antibodies produced by thehybridomas designated by Patent Deposit Designation PTA-10431 and PatentDeposit Designation PTA-10432 were each generated by immunizing anon-human mammal with a recombinant human soluble ST2 isolated fromhuman embryonic kidney (HEK)-293 cells.

Kits for measuring soluble ST2 are also commercially available, e.g.,the ST2 ELISA Kit manufactured by Medical & Biological Laboratories Co.,Ltd. (MBL International Corp., Woburn, Mass.), No. 7638. In addition,devices for measuring ST2 and other biomarkers are described in U.S.Patent Application Publication No. 2005/0250156 (incorporated herein byreference in its entirety).

As described in detail herein, soluble ST2 levels can be determined inany biological sample from a subject, including blood, serum, plasma,urine, and body tissue. Generally, the level of soluble ST2 isdetermined in a sample containing serum, blood, or plasma. The level ofsoluble ST2 can be determined in a biological sample that has beenstored for a period of time (e.g., for at least 1 hour, 1 day, 1 week,or 1 month) at a temperature at or below 10° C. (e.g., below 0° C.,below −20° C., or around −196° C.).

Elevated concentrations of soluble ST2 are markedly prognostic for deathin patients with heart failure, with a dramatic divergence in survivalcurves for those with elevated soluble ST2 soon after presentation(Weinberg et al., Circulation 107:721-726, 2003; Mueller et al., ClinChem. 54(4):752-756, 2008; Daniels et al., Am. Heart J. 160:721-728,2010; Ky et al., Circ. Heart Fail. 4(2):180-187, 2011; andManzano-Fernandez et al., Am. J. Cardiol. 107:259-267, 2011). Therelationship between soluble ST2 and death in heart failure patients wasshown to be independent of etiology, and superseded all other biomarkerpredictors of mortality in this setting, including other markers ofinflammation, myonecrosis, renal dysfunction, and most notablyNT-proBNP, a marker well known as having value for predicting death inheart failure patients.

Reference Levels of ST2

As described herein, the level of soluble ST2 in a subject indicateswhether a subject having a cardiovascular disease should be selected forexercise therapy, whether a subject will benefit or will not benefitfrom exercise therapy, or whether a subject should begin, continue, ordiscontinue exercise therapy. In addition, the level of soluble ST2 in asubject can be used to select a therapy including exercise for a subjecthaving a cardiovascular disease. Additional clinical and therapeuticuses of detecting a level of soluble ST2 are described herein and areknown in the art. Reference levels of human soluble ST2 can bedetermined using methods known in the art (e.g., using the antibodiesdescribed in U.S. patent application Ser. No. 13/083,333 and PCTApplication No. PCT/US2011/031801). In general, it will be desirable touse a reference level of soluble ST2 determined using the same method asis used to determine the level of soluble ST2 in the subject.

Risk Reference Levels

The methods described herein can include comparing the level of solubleST2 in a biological sample to a reference level of soluble ST2. Areference level of soluble ST2 can be or represent a level of solubleST2 found in a biological sample from a subject (e.g., a control subjectwho experienced or was more likely to experience an adverse outcome andengaged in exercise) or a population (e.g., a population of subjects whoexperienced or were more likely to experience an adverse outcome andengaged in exercise), or can be or represent a threshold level ofsoluble ST2 above which the risk of an adverse outcome is increased inthose who engange in exercise therapy. These reference levels arereferred to herein as the “risk reference level.” In general, thepresence of a level of ST2 below the risk reference level indicates thatthe subject does not have an elevated risk of an adverse outcome if theyengage in exercise therapy, while the presence of a level of ST2 abovethe risk reference level indicates that the subject has an elevated riskof an adverse outcome if they do engage in exercise therapy.

In some embodiments, the risk reference level of soluble ST2 is athreshold level of soluble ST2 or a percentile (e.g., 75th, 80th, 85th,90th, or 95th percentile) of soluble ST2 levels in a population ofsubjects who experienced an adverse outcome and engaged in exercise.

In some embodiments, efficacy reference level is about 55 ng/mL,determined using the Presage ST2 kit, or the equivalent thereof.

Efficacy Reference Levels

Alternatively or in addition, the methods described herein can includecomparing the level of soluble ST2 in a biological sample to a referencelevel of soluble ST2 that is or represents a level of soluble ST2 foundin a biological sample from a subject (e.g., a control subject whoexperienced a therapeutic benefit from exercise therapy, i.e., did notor was less likely to experience an adverse outcome and engaged inexercise) or a population (e.g., a population of subjects whoexperienced a therapeutic benefit from exercise therapy, i.e., did notexperience or were less likely to experience an adverse outcome andengaged in exercise), or can be or represent a threshold level ofsoluble ST2 below which the subject is likely to experience atherapeutic benefit from exercise therapy, i.e., a threshold level ofsoluble ST2 below which the likelihood of an adverse outcome isdecreased in those who engange in exercise therapy. These referencelevels are referred to herein as the “efficacy reference level.” Ingeneral, the presence of a level of ST2 below the efficacy referencelevel indicates that the subject has an increased likelihood of notexperiencing an adverse outcome if they engage in exercise therapy,while the presence of a level of ST2 above the efficacy reference levelindicates that the subject has an elevated risk of an adverse outcome ifthey do engage in exercise therapy.

In some embodiments, the efficacy reference level of soluble ST2 is athreshold level of soluble ST2 or a percentile (e.g., 75th, 80th, 85th,90th, or 95th percentile) of soluble ST2 levels in a population ofsubjects who experienced an adverse outcome and engaged in exercise.

In some embodiments, efficacy reference level is about 35 ng/mL,determined using the Presage ST2 kit, or the equivalent thereof.

Healthy Controls and Other Reference Levels

In some embodiments, the methods described herein can also includecomparing the level of soluble ST2 in a biological sample to a referencelevel of soluble ST2 that represent the average level of soluble ST2present in a population of subjects: a population of subjects diagnosedas having a specific cardiovascular disease, a population of healthysubjects not diagnosed with a disease (e.g., a healthy male patientpopulation or a healthy female patient population), a population ofsubjects not at risk of developing a cardiovascular disease, or apopulation of subjects not presenting with two or more symptoms of acardiovascular disease. A reference level can also be a baseline levelor a level in the same patient measured at an earlier or later point intime. Additional non-limiting examples of reference levels of humansoluble ST2 include the level of human soluble ST2 in a subject or apatient population that: does not have high risk cardiovascular disease;does not have renal failure; does not have hypertriglyceridemia,hypercholesterolemia, hypertension, and/or a body mass index of <30(e.g., a BMI under 25); and/or does not suffer from a pulmonary disease,sepsis, or Kawasaki disease.

In some embodiments, the reference level of soluble ST2 is a thresholdlevel of soluble ST2. In some embodiments, the threshold level ofsoluble ST2 is a median level of soluble ST2 or a percentile (e.g.,75th, 80th, 85th, 90th, or 95th percentile) of soluble ST2 levels in ahealthy patient population, e.g., a healthy male patient population or ahealthy female patient population (e.g., any of the values or rangeslisted in Table 1).

TABLE 1 sST2 Concentrations at Specific Thresholds - US Self-ReportedHealthy Cohort¹ Entire Cohort Male Female ST2 ST2 ST2 Percentiles(ng/mL) 95% CI (ng/mL) 95% CI (ng/mL) 95% CI 2.5 8.0 7.1 to 8.6 8.6  7.7to 11.8 7.3 5.5 to 8.4 5 9.3  8.4 to 10.2 11.8  8.6 to 12.7 8.5 7.3 to9.4 10 11.5 10.3 to 11.9 13.7 12.2 to 14.8 10.2  9.0 to 11.2 25 14.513.7 to 15.2 17.6 16.8 to 18.7 12.4 11.9 to 13.5 median 18.8 18.2 to19.9 23.6 21.3 to 25.1 16.2 15.4 to 17.4 75 25.3 23.8 to 26.9 30.6 28.7to 33.3 19.9 18.8 to 20.8 90 34.3 32.4 to 35.6 37.2 35.5 to 40.9 23.722.2 to 25.8 95 37.9 35.9 to 41.3 45.4 39.4 to 48.6 29.0 24.6 to 33.297.5 45.6 40.1 to 48.7 48.5 45.8 to 58.5 33.1 29.6 to 39.9 ¹These levelswere determined using the antibodies described in U.S. patentapplication Ser. No. 13/083,333 and PCT Application No.PCT/US2011/031801.

In some embodiments, the threshold level is 28.6 ng/mL, about 28 ng/mLto about 35 ng/mL, 35 ng/mL, about 35 ng/mL to about 45 ng/mL, about 45ng/mL to about 55 ng/mL, or the range of 55 to 60 ng/mL (e.g., theentire range or any level between 55 to 60 ng/mL) (e.g., using theantibodies described in U.S. patent application Ser. No. 13/083,333 andPCT Application No. PCT/US2011/031801). Reference levels of humansoluble ST2 can be determined using methods known in the art (e.g.,using the antibodies described in U.S. patent application Ser. No.13/083,333 and PCT Application No. PCT/US2011/031801). Additionalreference levels of soluble ST2 are known in the art. As is known in theart, the reference level of soluble ST2 can vary based on the assay usedto determine soluble ST2 levels.

Subjects

The methods described herein can be performed on a variety of subjectshaving a cardiovascular disease (e.g., any of the subjects describedherein). In some embodiments of any of the methods, the subject has beenpreviously diagnosed as having a cardiovascular disease. In someembodiments of any of the methods, the subject is hyperchlosterolemic,hypertriglyceridemic, hyperlidemic, a smoker, hypertensive, or has abody mass index of greater than 25 (e.g., between 25 and 30, or greaterthan 30). In some embodiments of any of the methods, the subject canalready be receiving a therapeutic agent (e.g., one or more of theadditional therapeutic agents described herein or known in the art fortreating a cardiovascular disease). In some embodiments, the subject hasheart failure.

In some embodiments of any of the methods, the subject can havepreviously been admitted to a hospital or can be receiving treatment onan outpatient basis. In some embodiments of any of the methods, thepatient can be 20 to 40 years old, 40 to 50 years old, 50 to 60 yearsold, 60 to 70 years old, 70 to 80 years old, 80 to 90 years old, or 90to 100 years old.

Methods of Treating a Subject Having a Cardiovascular Disease

Provided herein are methods of treating a subject having acardiovascular disease (e.g., any of the cardiovascular diseasesdescribed herein or known in the art, e.g., heart failure). Thesemethods include determining a level of soluble ST2 in a biologicalsample from the subject, identifying a subject that has a decreased(e.g., a significant or detectable decrease) level of soluble ST2 in thebiological sample compared to a risk or efficacy reference level ofsoluble ST2 (e.g., as described herein), and selecting the identifiedsubject for exercise therapy (e.g., any form of exercise therapydescribed herein or known in the art). Also provided are methods oftreating a subject that include determining a level of soluble ST2 in abiological sample from the subject, identifying a subject that has anincreased (e.g., a significant or detectable increase) level of solubleST2 in the biological sample compared to a risk or efficacy referencelevel of soluble ST2 (e.g., as described herein), and instructing theidentified subject not to begin, to discontinue, or to avoid exercisetherapy. The level of soluble ST2 in the biological sample can bedetermined using any of the methods described herein (e.g., methodsusing an antibody or antibody fragment that binds specifically tosoluble ST2). The biological sample can be any of the biological samplesdescribed herein. In some embodiments, the biological sample iscollected from a subject within 2 or 4 years of diagnosis with acardiovascular disease, a myocardial infarction, or heart failure.

The selected subject can begin performance of any of the exercisetherapy regimes described herein. Some embodiments of these methodsfurther include monitoring the subject to determine whether the exercisetherapy regime should be continued or altered in one or more aspects(e.g., any of the aspects of exercise therapy regimes described herein).Methods for monitoring the subject to determine whether the exercisetherapy should be continued or discontinued are described herein. Thesemethods can be performed by any health care professional (e.g., aphysician, a physical therapist, a nurse, a physician's assistant, alaboratory technician, or a nursing assistant).

In some embodiments, the treating results in a reduced (e.g., asignificant decrease) in the risk of an adverse outcome (e.g., any ofthe adverse outcomes described herein) in the subject. In someembodiments, the treating results in a decrease in the number ofsymptoms of a cardiovascular disease, a decrease (e.g., a significant ordetectable decrease) in the intensity, frequency, or duration of one ormore symptoms of a cardiovascular disease, or a decrease in the level ofat least one marker of a cardiovascular disease in a biological samplefrom the subject.

Methods of Selecting a Therapy for a Subject Having a CardiovascularDisease

Also provided are methods of selecting a therapy for a subject having acardiovascular disease. These methods include determining a level ofsoluble ST2 in a biological sample from the subject and comparing thelevel of soluble ST2 in the biological sample to a risk or efficacyreference level of soluble ST2 (e.g., any of the reference levels ofsoluble ST2 described herein or known in the art), where a decreased(e.g., a significant or detectable decrease) level of soluble ST2 in thebiological sample compared to the risk or efficacy reference level ofsoluble ST2 indicates that the subject should begin or continue exercisetherapy (e.g., any of the exercise therapy regimes described herein),and an elevated (e.g., a significant or detectable increase) level ofsoluble ST2 in the biological sample compared to the risk or efficacyreference level of soluble ST2 indicates that the subject should notbegin or should discontinue exercise therapy. The level of soluble ST2in the biological sample can be determined using any of the methodsdescribed herein, and the biological sample can be any of the biologicalsamples described herein. In some embodiments, the biological sample iscollected from a subject within 2 or 4 years of diagnosis with acardiovascular disease, a myocardial infarction, or heart failure. Thesemethods can be performed by any health care professional (e.g., aphysician, a physical therapist, a nurse, a physician's assistant, alaboratory technician, or a nursing assistant). In some embodiments, thesubject can already be performing an exercise therapy regime.

Methods of Identifying a Subject that Will Benefit/not Benefit fromExercise Therapy

Also provided herein are methods for identifying a subject that willbenefit from exercise therapy (benefit from the performance of any ofthe exercise therapy regimes described herein). The methods includedetermining a level of soluble ST2 in a biological sample from thesubject, and selecting a subject that has a decreased (e.g., asignificant or detectable decrease) level of soluble ST2 in thebiological sample compared to an efficacy reference level of soluble ST2(e.g., any of the reference levels of soluble ST2 described herein orknown in the art), where the selected subject is identified as a subjectthat will benefit from exercise therapy (e.g., any of the exercisetherapy regimes described herein). Also provided are methods ofidentifying a subject that will not benefit from exercise therapy (willnot benefit from the performance of any of the exercise therapy regimesdescribed herein) that include determining a level of soluble ST2 in abiological sample from the subject, and selecting a subject that has anelevated (e.g., a significant or detectable increase) level of solubleST2 in the biological sample compared to an efficacy reference level ofsoluble ST2 (e.g., any of the reference levels of soluble ST2 describedhere in or known in the art), where the selected subject is identifiedas a subject that will not benefit from exercise therapy (e.g., any ofthe exercise therapy regimes described herein).

In some embodiments, the biological sample is collected from a subjectwithin 2 or 4 years of diagnosis with a cardiovascular disease, amyocardial infarction, or heart failure. In some embodiments, theefficacy reference level of soluble ST2 is a threshold soluble ST2 levelof less than or equal to 35 ng/mL or a range of 55 to 60 ng/mL (e.g.,the entire range or any level between 55 to 60 ng/mL). The level ofsoluble ST2 in the biological sample can be determined using any of themethods described herein (e.g., methods using an antibody or antibodyfragment that binds specifically to soluble ST2), and the biologicalsample can be any of the biological samples described herein.

In some embodiments of these methods, the subject has been previouslydiagnosed as having a cardiovascular disease (e.g., previously diagnosedas having heart failure). In some embodiments, the subject ishyperchlosterolemic, hypertriglyceridemic, hyperlidemic, a smoker,hypertensive, or has a body mass index of greater than 25 (e.g., between25 and 30, or greater than 30). In some embodiments, the subject canalready be receiving a therapeutic agent (e.g., one or more of theadditional therapeutic agents described herein or known in the art fortreating a cardiovascular disease). In some embodiments, the subject canhave previously been admitted to a hospital or can be receivingtreatment on an outpatient basis. In some embodiments, the patient canbe 20 to 40 years old, 40 to 50 years old, 50 to 60 years old, 60 to 70years old, 70 to 80 years old, 80 to 90 years old, or 90 to 100 yearsold.

In some embodiments, the benefit from exercise therapy can be one ormore of the following: a reduction (e.g., a significant decrease) in therisk of an adverse outcome (e.g., any of the adverse outcomes describedherein) in the subject, a reduction in the number of symptoms of acardiovascular disease, a reduction (e.g., a detectable or observabledecrease) in the intensity, frequency, or duration of one or moresymptoms of a cardiovascular disease, or a reduction (e.g., detectabledecrease) in the levels of at least one marker of a cardiovasculardisease in a biological sample from the subject (e.g., as compared to asubject or population of subjects having the same cardiovascular diseasebut not receiving therapy or receiving a different therapy). The benefitfrom exercise therapy can be determined at various time points in asubject (e.g., after at least 6 months of exercise therapy, after 1 yearof exercise therapy, or after 2 years of exercise therapy).

Methods of Determining Whether a Subject should Begin, Continue, orDiscontinue Exercise Therapy

Also provided are methods of determining whether a subject having acardiovascular disease should begin or continue exercise therapy (e.g.,any of the exercise therapy regimes described herein) that includedetermining a level of soluble ST2 in a biological sample from thesubject, wherein a decreased (e.g., a significant or detectabledecrease) level of soluble ST2 in the biological sample compared to arisk or efficacy reference level of soluble ST2 (e.g., any of thereference levels described herein) indicates that the subject shouldbegin or continue exercise therapy. Also provided are methods ofdetermining whether a subject having a cardiovascular disease should notbegin or discontinue exercise therapy (e.g., any of the exercise therapyregimes described herein) that include determining a level of solubleST2 in a biological sample from the subject, where an elevated (e.g., asignificant or detectable increase) level of soluble ST2 in thebiological sample compared to a risk or efficacy reference level ofsoluble ST2 (e.g., any of the reference levels of soluble ST2 describedherein) indicates that the subject should not begin or discontinueexercise therapy.

Also provided are methods of determining whether a subject having acardiovascular disorder should discontinue or avoid exercise therapy(e.g., any of the exercise therapy regimes described herein) thatinclude determining a level of soluble ST2 in a biological sample fromthe subject at a first time point before or after the start of exercisetherapy, and determining a level of soluble ST2 in a biological samplefrom the subject undergoing (performing) exercise therapy at a secondtime point after the start of exercise therapy and after the first timepoint, where an increase (e.g., a significant or detectable increase) inthe level of soluble ST2 in the biological sample of the second timepoint compared to the level of soluble ST2 in the biological sample atthe first time point indicates that the subject should discontinue oravoid exercise therapy. Also provided are methods of determining whethera subject having a cardiovascular disorder should continue exercisetherapy (e.g., any of the exercise therapy regimes described herein)that include determining a level of soluble ST2 in a biological samplefrom the subject at a first time point before or after the start ofexercise therapy, and determining a level of soluble ST2 in a biologicalsample from the subject undergoing (performing) exercise therapy at asecond time point after the start of exercise therapy and after thefirst time point, where a decrease (e.g., a significant or detectabledecrease) or no change (e.g., no significant change) in the level ofsoluble ST2 in the biological sample of the second time point comparedto the level of soluble ST2 in the biological sample at the first timepoint indicates that the subject should continue exercise therapy.

Also provided are methods of determining whether a subject having acardiovascular disorder should continue exercise therapy (e.g., any ofthe exercise therapy regimes described herein) that include determininga level of soluble ST2 in a biological sample from the subject at afirst time point before or after the start of exercise therapy, andinstructing the subject to begin or continue exercise therapy if thelevel of soluble ST2 is below a risk or efficacy reference level; anddetermining a level of soluble ST2 in a biological sample from thesubject undergoing (performing) exercise therapy at a second time pointafter the start of exercise therapy and after the first time point, andinstructing the subject to continue the exercise therapy if the level atthe second time point is still below the risk or efficacy referencelevel, or to stop the exercise therapy if the level at the second timepoint is aboce the risk or efficacy reference level.

In some embodiments, one or more additional levels of soluble ST2 can bedetermined in the subject (e.g., determined in biological samplesobtained at one or more additional time points after the second timepoint). In some embodiments, the level of soluble ST2 is determined in abiological sample obtained from the subject having cardiovasculardisease at least every month (e.g., at least every two months, at leastevery three months, at least every four months, at least every fivemonths, or at least every six months) during the performance of aexercise therapy regime. In such embodiments, an increase (e.g., asignificant or detectable increase) in the level of soluble ST2 in abiological sample taken at a later time point compared to the level ofsoluble ST2 in a biological sample taken at an earlier time point (e.g.,the immediately prior biological sample), or the presence of a levelabove a risk or efficacy reference level, indicates that the subjectshould discontinue exercise therapy, and a decrease (e.g., a significantor detectable decrease) or no change (e.g., no significant change) inthe level of soluble ST2 in a biological sample taken at a later timepoint compared to the level of soluble ST2 in a biological sample takenat an earlier time point (e.g., the immediately prior biologicalsample), or the presence of a level below a risk or efficacy referencelevel, indicates that the subject should continue exercise therapy. Insome embodiments of these methods, the subjects are monitored by ahealth care professional (e.g., a physician, a physical therapist, anurse, a nurse's assistant, a physician's assistant, or a laboratorytechnician).

In some embodiments, the biological sample is collected from a subjectwithin 2 years of diagnosis with a cardiovascular disease, a myocardialinfarction, or heart failure. The level of soluble ST2 in the biologicalsample can be determined using any of the methods described herein, andthe biological sample can be any of the biological samples describedherein. These methods can reduce (e.g., a significant reduction) in therisk of death or an adverse outcome (e.g., any of the adverse outcomesdescribed herein) in the subject, reduce the number of symptoms of acardiovascular disease, reduce (e.g., a detectable or observablereduction) in the intensity, frequency, or duration in one or moresymptoms of a cardiovascular disease, or reduce (e.g., detectablereduction) in the level of at least one marker (e.g., any of the markersdescribed herein) of a cardiovascular disease in a biological samplefrom the subject (e.g., as compared to a subject or population ofsubjects having the same cardiovascular disease but not receivingtherapy or receiving a different therapy).

Methods of Reducing the Risk of an Adverse Outcome in a Subject

Also provided herein are methods of reducing (e.g., a significantreduction) the risk of an adverse outcome (e.g., risk of death) in asubject having a cardiovascular disease that include determining a levelof soluble ST2 in a biological sample from the subject, identifying asubject that has a decreased (e.g., a significant or detectabledecrease) level of soluble ST2 in the biological sample compared to arisk reference level of soluble ST2 (e.g., as described herein), andselecting the identified subject for exercise therapy (e.g., any of theexercise therapy regimes described herein). Also provided are methods ofreducing (e.g., a significant reduction) the risk of an adverse outcome(e.g., risk of death) in a subject having a cardiovascular disease thatinclude determining a level of soluble ST2 in a biological sample fromthe subject, identifying a subject that has an elevated (e.g., asignificant or detectable increase) level of soluble ST2 in thebiological sample compared to a risk reference level of soluble ST2(e.g., as described herein), and instructing the identified subject toavoid or discontinue exercise therapy (e.g., any of the exercise therapyregimes described herein).

In some embodiments, the adverse outcome can be one or more of: death,organ failure, organ transplantation, hospitalization orrehospitalization, recurrence of one or more symptoms of acardiovascular disease, development of one or more additional symptomsof a cardiovascular disease, an increase in the frequency, intensity, orduration of one or more symptoms of a cardiovascular disease experiencedby the subject, or a first or subsequent myocardial infarction. In someembodiments, the biological sample is collected from a subject within 2years of diagnosis with a cardiovascular disease, a myocardialinfarction, or heart failure. Any of these methods can be performed by ahealth care professional (e.g., a physician, a physical therapist, anurse, a nurse's assistant, a physician's assistant, or a laboratorytechnician).

Methods of Predicting the Efficacy of Exercise Therapy

Also provided are methods of predicting the efficacy of exercise therapy(e.g., any of the exercise therapy regimes described herein) in asubject having a cardiovascular disease. These methods includedetermining a level of soluble ST2 in a biological sample from thesubject, and comparing the level of soluble ST2 in the biological sampleto an efficacy reference level of soluble ST2 (e.g., any of thereference levels of soluble ST2 described herein), wherein a decreased(e.g., a significant or detectable decrease) level of soluble ST2 in thebiological sample compared to the efficacy reference level of solubleST2 (e.g., as described herein) indicates that exercise therapy will beeffective in the subject, and an elevated (e.g., a significant ordetectable increase) level of soluble ST2 in the biological samplecompared to the efficacy reference level of soluble ST2 (e.g., asdescribed herein) indicates that the exercise therapy will not beeffective in the subject. In some embodiments, the efficacy referencelevel of soluble ST2 is a threshold soluble ST2 level of 28.6 ng/mL,less than or equal to 35 ng/mL, about 28 ng/mL to about 35 ng/mL (e.g.,the entire range or any level between 28 to 35 ng/mL), 35 ng/mL, about35 ng/mL to about 45 ng/mL (e.g., the entire range or any level between35 to 45 ng/mL), about 45 ng/mL to about 55 ng/mL (e.g., the entirerange or any level between 45 to 55 ng/mL), about 35 ng/mL to about 55ng/mL (e.g., the entire range or any level between 35 to 55 ng/mL), orthe range of 55 to 60 ng/mL (e.g., the entire range or any level between55 to 60 ng/mL) (e.g., determined using the antibodies described in U.S.patent application Ser. No. 13/083,333 and PCT Application No.PCT/US2011/031801). In some embodiments, the biological sample iscollected from a subject within 2 years of diagnosis with acardiovascular disease, a myocardial infarction, or heart failure. Thelevel of soluble ST2 in the biological sample can be determined usingany of the methods described herein, and the biological sample can beany of the biological samples described herein.

In some embodiments, the efficacy of exercise therapy can be one or moreof the following: a reduction (e.g., a significant decrease) in the riskof an adverse outcome (e.g., any of the adverse outcomes describedherein) in the subject, a reduction in the number of symptoms of acardiovascular disease, a reduction (e.g., a detectable or observabledecrease) in the intensity, frequency, or duration of one or moresymptoms of a cardiovascular disease, or a reduction (e.g., detectabledecrease) in the levels of at least one marker of a cardiovasculardisease (e.g., any of the markers described herein or known in the art)in a biological sample from the subject (e.g., as compared to a subjector population of subjects having the same cardiovascular disease but notreceiving therapy or receiving a different therapy). The efficacy ofexercise therapy can be determined at various time points in a subject(e.g., after at least 6 months of exercise therapy, after 1 year ofexercise therapy, or after 2 years of exercise therapy).

Additional Therapeutic Markers

Any of the methods described herein can further include determining thelevel of at least one additional marker (e.g., at least one additionalmarker of a cardiovascular disease) in a biological sample from asubject. In some embodiments, the biological sample used to determinethe level of the at least one additional marker may be the samesample(s) that is used to determine a level of soluble ST2 in a subject.In some embodiments, the biological sample used to determine the levelof the at least one additional marker is a different sample than thesample(s) used to determine a level of soluble ST2 in a subject. Thebiological sample can be any of the biological samples described herein.

The additional marker can be any protein, nucleic acid, lipid, orcarbohydrate, or a combination (e.g., two or more) thereof, that isdiagnostic of the presence of a particular disease (e.g., diagnostic ofa cardiovascular disease). Several additional markers useful for thediagnosis of a cardiovascular disease are known in the art, and include,without limitation, cardiac troponin I, B-type natriuretic peptide(e.g., proBNP, NT-proBNP, and BNP), atrial natriuretic peptide (e.g.,proANP, NT-proANP, and ANP), troponin, C-reactive protein, creatinine,Blood Urea Nitrogen (BUN), liver function enzymes, albumin, andbacterial endotoxin. Additional non-limiting markers of a cardiovasculardisease are described in U.S. Patent Application Publication Nos.:2007/0248981; 2011/0053170; 2010/0009356; 2010/0055683; and 2009/0264779(each of which is hereby incorporated by reference). Additional markersof a cardiovascular disease are known in the art.

Methods for determining the level of the above described markers of acardiovascular disease are known in the art. Diagnostic tests fordetermining the level of several of these markers are commerciallyavailable. For example, diagnostic tests for determining the level ofC-reactive protein (e.g., Exocell), B-type natriuretic peptide (e.g.,Alpco Immunoassays), atrial natriuretic peptide (e.g., Cusabio BiotechCo., Ltd.), and cardiac troponin I (e.g., Calbiotech Inc.) arecommercially available.

Additional Therapeutic Treatments

In any of the methods described herein, the subject can further beadministered an additional therapeutic treatment (e.g., at least onetherapeutic treatment in addition to exercise therapy). In someembodiments of the methods described herein, the subject can bereceiving at least one therapeutic treatment at the time the exercisetherapy begins. In some embodiments, a health care professional mayadjust (e.g., increase or decrease) the dosage or frequency ofadministration of at least one therapeutic agent administered to thesubject prior to the start of exercise therapy or at a time point duringthe exercise therapy. In some embodiments of the methods describedherein, the efficacy of the exercise therapy allows for a decrease inthe number of therapeutic agents or allows for a decrease in the dose orfrequency of administration of one or more therapeutic agents to asubject having a cardiovascular disease.

Non-limiting examples of therapeutic treatment of a cardiovasculardisease (in addition to exercise therapy) include the administration ofone of more of the following agents: statins, anti-inflammatory agents,anti-thrombotic agents, anti-coagulants, anti-platelet agents,lipid-reducing agents, direct thrombin inhibitors, glycoprotein IIb/IIIbreceptor inhibitors, calcium channel blockers, beta-adrenergic receptorblockers, cyclooxygenase-2 inhibitors, and renin-angiotensin-aldosteronesystem (RAAS) inhibitors.

Non-limiting examples of lipid-reducing agents that can be used to treata cardiovascular disease in a subject (alone or in combination any othertherapy, including exercise therapy) include: a statin, gemfibrozil,cholystyramine, colestipol, nicotinic acid, and probucol. Statins aremolecules that are capable of inhibiting the activity of HMG-CoAreductase. Non-limiting examples of statins that can be administered toa subject having a cardiovascular disease (alone or in combination withany other therapy, including exercise therapy) include: atorvastatin,cirivastatin, fluvastatin, lovastatin, pitavastatin, pravastatin,rosuvastatin, and simvastatin. Additional examples of statins and otherlipid-reducing agents are known in the art.

Non-limiting examples of anti-inflammatory agents that can be used totreat a cardiovascular disease in a subject (alone or in combination anyother therapy, including exercise therapy) include: Alclofenac,Alclometasone Dipropionate, Algestone Acetonide, Alpha Amylase,Amcinafal, Amcinafide, Amfenac Sodium, Amiprilose Hydrochloride,Anakinra, Anirolac, Anitrazafen, Apazone, Balsalazide Disodium,Bendazac, Benoxaprofen, Benzydamine Hydrochloride, Bromelains,Broperamole, Budesonide, Carprofen, Cicloprofen, Cintazone, Cliprofen,Clobetasol Propionate, Clobetasone Butyrate, Clopirac, CloticasonePropionate, Cormethasone Acetate, Cortodoxone, Deflazacort, Desonide,Desoximetasone, Dexamethasone Dipropionate, Diclofenac Potassium,Diclofenac Sodium, Diflorasone Diacetate, Diflumidone Sodium,Diflunisal, Difluprednate, Diftalone, Dimethyl Sulfoxide, Drocinonide,Endrysone, Enlimomab, Enolicam Sodium, Epirizole, Etodolac, Etofenamate,Felbinac, Fenamole, Fenbufen, Fenclofenac, Fenclorac, Fendosal,Fenpipalone, Fentiazac, Flazalone, Fluazacort, Flufenamic Acid,Flumizole, Flunisolide Acetate, Flunixin, Flunixin Meglumine, FluocortinButyl, Fluorometholone Acetate, Fluquazone, Flurbiprofen, Fluretofen,Fluticasone Propionate, Furaprofen, Furobufen, Halcinonide, HalobetasolPropionate, Halopredone Acetate, Ibufenac, Ibuprofen, IbuprofenAluminum, Ibuprofen Piconol, Ilonidap, Indomethacin, IndomethacinSodium, Indoprofen, Indoxole, Intrazole, Isoflupredone Acetate,Isoxepac, Isoxicam, Ketoprofen, Lofemizole Hydrochloride, Lornoxicam,Loteprednol Etabonate, Meclofenamate Sodium, Meclofenamic Acid,Meclorisone Dibutyrate, Mefenamic Acid, Mesalamine, Meseclazone,Methylprednisolone Suleptanate, Morniflumate, Nabumetone, Naproxen,Naproxen Sodium, Naproxol, Nimazone, Olsalazine Sodium, Orgotein,Orpanoxin, Oxaprozin, Oxyphenbutazone, Paranyline Hydrochloride,Pentosan Polysulfate Sodium, Phenbutazone Sodium Glycerate, Pirfenidone,Piroxicam, Piroxicam Cinnamate, Piroxicam Olamine, Pirprofen,Prednazate, Prifelone, Prodolic Acid, Proquazone, Proxazole, ProxazoleCitrate, Rimexolone, Romazarit, Salcolex, Salnacedin, Salsalate,Salycilates, Sanguinarium Chloride, Seclazone, Sermetacin, Sudoxicam,Sulindac, Suprofen, Talmetacin, Talniflumate, Talosalate, Tebufelone,Tenidap, Tenidap Sodium, Tenoxicam, Tesicam, Tesimide, Tetrydamine,Tiopinac, Tixocortol Pivalate, Tolmetin, Tolmetin Sodium, Triclonide,Triflumidate, Zidometacin, Glucocorticoids, and Zomepirac Sodium. Onepreferred anti-inflammatory agent is aspirin. Additional examples ofanti-inflammatory agents are known in the art.

Non-limiting examples of anti-thrombotic agents that can be used totreat a cardiovascular disease in a subject (alone or in combinationwith any other therapy, including exercise therapy) include: plasminogenproactivator, tissue plasminogen activator, AnisoylatedPlasminogen-Streptokinase Activator Complex, Pro-Urokinase, (Pro-UK),rTPA (recombinant alteplase or activase), recombinant Pro-UK,Abbokinase, Eminase, Sreptase Anagrelide Hydrochloride, Bivalirudin,Dalteparin Sodium, Danaparoid Sodium, Dazoxiben Hydrochloride, EfegatranSulfate, Enoxaparin Sodium, Ifetroban, Ifetroban Sodium, TinzaparinSodium, Retaplase, Trifenagrel, Warfarin, and Dextrans. Additionalexamples of anti-thrombotic agents are known in the art.

Non-limiting examples of anti-coagulants that can be used to treat acardiovascular disease in a subject (alone or in combination with anyother therapy, including exercise therapy) include: Ancrod,Anticoagulant Citrate Dextrose Solution, Anticoagulant Citrate PhosphateDextrose Adenine Solution, Anticoagulant Citrate Phosphate DextroseSolution, Anticoagulant Heparin Solution, Anticoagulant Sodium CitrateSolution, Ardeparin Sodium, Bivalirudin, Bromindione, Dalteparin Sodium,Desirudin, Dicumarol, Heparin Calcium, Heparin Sodium, Lyapolate Sodium,Nafamostat Mesylate, Phenprocoumon, Tinzaparin Sodium, and WarfarinSodium. Additional examples of anti-coagulants are known in the art.

Non-limiting examples of anti-platelet agents that can be used to treata cardiovascular disease in a subject (alone or in combination with anyother therapy, including exercise therapy) include: Clopridogrel,Sulfinpyrazone, Aspirin, Dipyridamole, Clofibrate, Pyridinol Carbamate,Prostaglandin E, Glucagon, Antiserotonin drugs, Caffeine, TheophyllinPentoxifyllin, Ticlopidine, and Anagrelide. Additional examples ofanti-platelet agents are known in the art.

Non-limiting examples of direct thrombin inhibitors that can be used totreat a cardiovascular disease in a subject (alone or in combinationwith any other therapy, including exercise therapy) include: hirudin,hirugen, hirulog, agatroban, PPACK, and thrombin aptamers. Additionalexamples of thrombin inhibitors are known in the art.

Non-limiting examples of glycoprotein IIb/IIIb receptor inhibitors thatcan be used to treat a cardiovascular disease in a subject (alone or incombination with any other therapy, including exercise therapy) include:ReoPro (abcixamab), lamifiban, and tirofiban. Additional examples ofglycoprotein IIb/IIIb receptor inhibitors are known in the art.

Non-limiting examples of calcium channel blockers that can be used totreat a cardiovascular disease in a subject (alone or in combinationwith any other therapy, including exercise therapy) include:dihydropyridines, such as nifedipine; phenyl alkyl amines, such asverapamil; and benzothiazepines, such as diltiazem. Additionalnon-limiting examples of calcium channel blockers include amrinone,amlodipine, bencyclane, felodipine, fendiline, flunarizine, isradipine,nicardipine, nimodipine, perhexilene, gallopamil, tiapamil, tiapamilanalogues (such as 1993RO-11-2933), phenytoin, barbiturates, and thepeptides dynorphin, omega-conotoxin, and omega-agatoxin, and the likeand/or pharmaceutically acceptable salts thereof. Additional examples ofcalcium channel blockers are known in the art.

Non-limiting examples of beta-adrenergic receptor blockers that can beused to treat a cardiovascular disease in a subject (alone or incombination with any other therapy, including exercise therapy) include:atenolol, acebutolol, alprenolol, befunolol, betaxolol, bunitrolol,carteolol, celiprolol, hedroxalol, indenolol, labetalol, levobunolol,mepindolol, methypranol, metindol, metoprolol, metrizoranolol,oxprenolol, pindolol, propranolol, practolol, practolol, sotalolnadolol,tiprenolol, tomalolol, timolol, bupranolol, penbutolol, trimepranol,2-(3-(1,1-dimethylethyl)-amino-2-hydroxypropoxy)-3-pyridenecarbonitrilHCl, 1-butylamino-3-(2,5-dichlorophenoxy)-2-propanol,1-isopropylamino-3-(4-(2-cyclopropylmethoxyethyl)phenoxy)-2-propanol,3-isopropylamino-1-(7-methylindan-4-yloxy)-2-butanol,2-(3-t-butylamino-2-hydroxy-propylthio)-4-(5-carbamoyl-2-thienyl)thiazol,and 7-(2-hydroxy-3-t-butylaminpropoxy)phthalide. The abovebeta-adrenergic receptor blockers can be used as isomeric mixtures, orin their respective levorotating or dextrorotating form. Additionalexamples of beta-adrenergic receptor blockers are known in the art.

Non-limiting examples of cyclooxygenase-2 inhibitors that can be used totreat a cardiovascular disease in a subject (alone or in combinationwith any other therapy, including exercise therapy) include thosedescribed in U.S. Pat. Nos. 5,474,995; 5,521,213; 5,536,752; 5,550,142;5,552,422; 5,604,253; 5,604,260; 5,639,780; 5,677,318; 5,691,374;5,698,584; 5,710,140; 5,733,909; 5,789,413; 5,817,700; 5,849,943;5,861,419; 5,922,742; 5,925,631; 5,643,933; 5,474,995; and 5,543,297; WO95/00501, and WO 95/18799 (each of which is incorporated herein byreference). Additional examples of cyclooxygenase-2 inhibitors are knownin the art.

Renin-angiotensin-aldosterone system (RAAS) inhibitors can be used totreat a cardiovascular disease in a subject (alone or in combinationwith any other therapy, such as exercise therapy). RAAS inhibitorsinclude agents that interfere with the function and synthesis orcatabolism of angiotensin II. RAAS agents include, but are not limitedto, angiotensin-converting enzyme (ACE) inhibitors, angiotensin IIreceptor blockers, agents that activate the catabolism of angiotensinII, agents that prevent the synthesis of angiotensin I (from whichangiotensin II is ultimately derived), and aldosterone antagonists. TheRAAS is involved in the regulation of hemodynamics and water andelectrolyte balance. Factors that lower blood volume, renal perfusionpressure, or the concentration of Na⁺ in plasma tend to activate thesystem, while factors that increase these parameters tend to suppressits function. RAAS inhibitors are compounds that act to interfere withthe production of angiotensin II from angiotensinogen or angiotensin Ior interfere with the activity of angiotensin II. Such inhibitors arewell known in the art and include compounds that act to inhibit theenzymes involved in the ultimate production of angiotensin II, includingrenin and ACE. They also include compounds that interfere with theactivity of angiotensin II, once produced.

Angiotensin II receptor blockers include angiotensin II antagonistswhich interfere with the activity of angiotensin II by binding toangiotensin II receptors and interfere with their activity. AngiotensinII receptor blockers are well known and include peptide compounds andnon-peptide compounds. Most angiotensin II receptor blockers areslightly modified congeners in which agonist activity is attenuated byreplacement of phenylalanine in position 8 with some other amino acid.Examples of angiotensin II receptor blockers include: peptidic compounds(e.g., saralasin, [(San1)(Val5)(Ala8)] angiotensin-(1-8) octapeptide,and related analogs); N-substituted imidazole-2-one (U.S. Pat. No.5,087,634); imidazole acetate derivatives, including2-N-butyl-4-chloro-1-(2-chlorobenzile), imidazole-5-acetic acid (see,Long et al., J. Pharmacol. Exp. Ther. 247:1-7, 1988); 4, 5, 6,7-tetrahydro-1H-imidazo [4, 5-c] pyridine-6-carboxylic acid, and analogderivatives thereof (U.S. Pat. No. 4,816,463); N2-tetrazolebeta-glucuronide analogs (U.S. Pat. No. 5,085,992); substitutedpyrroles, pyrazoles, and tryazoles (U.S. Pat. No. 5,081,127); phenol andheterocyclic derivatives, such as 1, 3-imidazoles (U.S. Pat. No.5,073,566); imidazo-fused 7-member ring heterocycles (U.S. Pat. No.5,064,825); peptides (e.g., U.S. Pat. No. 4,772,684); antibodies toangiotensin II (e.g., U.S. Pat. No. 4,302,386); and aralkyl imidazolecompounds, such as biphenyl-methyl substituted imidazoles (e.g., EP253,310); N-morpholinoacetyl-(-1-naphthyl)-L-alanyl-(4,thiazolyl)-L-alanyl (35,45)-4-amino-3-hydroxy-5-cyclo-hexapentanoyl-N-hexylamide; SKF108566(E-alpha-2-[2-butyl-1-(carboxyphenyl)methyl]1H-imidazole-5-yl[methylane]-2-thiophenepropanoic acid); Losartan;Remikirin; and A2 agonists.

Non-limiting examples of ACE inhibitors include acylmercapto andmercaptoalkanoyl prolines, such as captopril (U.S. Pat. No. 4,105,776)and zofenopril (U.S. Pat. No. 4,316,906); carboxyalkyl dipeptides, suchas enalapril (U.S. Pat. No. 4,374,829), lisinopril (U.S. Pat. No.4,374,829), quinapril (U.S. Pat. No. 4,344,949), ramipril (U.S. Pat. No.4,587,258), and perindopril (U.S. Pat. No. 4,508,729); carboxyalkyldipeptide mimics, such as cilazapril (U.S. Pat. No. 4,512,924) andbenazapril (U.S. Pat. No. 4,410,520); and phosphinylalkanoyl prolines,such as fosinopril (U.S. Pat. No. 4,337,201) and trandolopril.

Additional non-limiting examples of RAAS inhibitors include: derivativesof peptides (U.S. Pat. No. 5,116,835); amino acids connected bynonpeptide bonds (U.S. Pat. No. 5,114,937); di- and tri-peptidederivatives (U.S. Pat. No. 5,106,835); amino acids and derivativesthereof (U.S. Pat. Nos. 5,104,869 and 5,095,119); diol sulfonamides andsulfinyls (U.S. Pat. No. 5,098,924); modified peptides (U.S. Pat. No.5,095,006); peptidyl beta-aminoacyl aminodiol carbamates (U.S. Pat. No.5,089,471); pyrolimidazolones (U.S. Pat. No. 5,075,451); fluorine andchlorine statine, or statone containing peptides (U.S. Pat. No.5,066,643); peptidyl amino diols (U.S. Pat. Nos. 5,063,208 and4,845,079); N-morpholino derivatives (U.S. Pat. No. 5,055,466);pepstatin derivatives (U.S. Pat. No. 4,980,283); N-heterocyclic alcohols(U.S. Pat. No. 4,885,292); monoclonal antibodies to renin (U.S. Pat. No.4,780,401); and a variety of other peptides and analogs thereof (U.S.Pat. Nos. 5,071,837, 5,064,965, 5,063,207, 5,036,054, 5,036,053,5,034,512, and 4,894,437) (each of which is incorporated by reference).

Additional examples of RAAS inhibitors include aldosterone antagonists.Non-limiting examples of aldosterone antagonists include:Spironolactone, Eplerenone, Canrenone (canrenoate potassium), Prorenone(prorenoate potassium), and Mexrenone (mexrenoate potassium).

EXAMPLES

The invention is further described in the following examples, which donot limit the scope of the invention described in the claims.

Example 1 Soluble ST2 Levels are Predictive of Mortality and AdverseEvents in Heart Failure Patients

Analysis was performed on the samples and data collected from 2331 heartfailure patients enrolled in the HF-ACTION study. A total of 2329 of thepatients underwent baseline exercise testing and were randomized toexercise therapy or normal clinical therapy. Of these patients, bloodsamples were collected from 912 patients representing both arms of thestudy, 453 were in the exercise arm and 459 were in the normal treatmentarm. Nine hundred and ten of these patients had a sufficient sample sizefor soluble ST2 measurement. The median soluble ST2 level in this cohortwas 23.7 ng/ml, and ranged from 2.2 ng/mL to 344.2 ng/ml. As in thepreviously published heart failure cohorts, soluble ST2 levels werepredictive of death for the full duration of the follow-up period (up to1460 days). The prognostic value of soluble ST2 levels for risk of deathin heart failure patients during the entire follow-up period (up to 1460days) is illustrated in FIG. 1. Patients in the highest soluble ST2level group (soluble ST2 levels of greater than 28.6 ng/mL) have thegreatest risk of mortality, with this risk presenting early in thefollow-up period. The increased risk of mortality for those patients inthe highest soluble ST2 level group is maintained throughout the entirefollow-up period.

The predictive strength of soluble ST2 levels was also assessed by Coxproportional hazards analysis, using soluble ST2 level as both a log(ln)-transformed continuous variable (HR 3.38, p<0.0001), as well as adichotomous variable at a concentration of 35 ng/ml (HR 2.59, p<0.0001).FIG. 2 shows the surviving portion of heart failure patients over timefor subjects having ST2 concentrations above or below a level of 35ng/mL.

Example 2 Subjects with Elevated Levels of Soluble ST2 Performing anExercise Therapy Regime have an Increased Risk of Death

As reported in O'Connor et al. (JAMA 301(14):1439-1450, 2009) there wasa modest, but insignificant, decrease in mortality rate in heart failurepatients who underwent exercise therapy relative to those who receivedstandard care. An assessment of soluble ST2 levels in this cohort showsthat the therapeutic efficacy of exercise therapy is different inpatients having a high versus low level of soluble ST2, with morebenefit, e.g., lower mortality rate, observed in heart failure patientswith low soluble ST2 levels. This correlation is illustrated in aKaplan-Meier analysis of the data from the following heart failurepatient groups: patients having a level of soluble ST2 less than orequal to 35 ng/mL and performing an exercise therapy regime (line 1, topline); patients having a level of soluble ST2 less than or equal to 35ng/nL and not performing an exercise therapy regime (line 2, second linefrom the top); patients having a level of soluble ST2 greater than 35ng/mL and not performing an exercise therapy regime (line 3, second linefrom the bottom); and patients having a level of soluble ST2 greaterthan 35 ng/mL and performing an exercise therapy regime (line 4, bottomline) (FIG. 3). In both treatment arms (i.e., those performing anexercise therapy regime or those receiving standard care (not performingan exercise therapy regime)), heart failure patients with low solubleST2 levels have better survival over the 4 year follow-up period thanpatients with elevated soluble ST2 levels. By 1 year, the patients withlow soluble ST2 levels who performed an exercise therapy regime had asignificantly lower mortality rate (˜3%) compared to the usual caretreatment group (not performing an exercise therapy regime), and thisbenefit persisted for the full 4-year follow-up period. Conversely, thedata from heart failure patients with elevated soluble ST2 levels showno survival benefit from the performance of an exercise therapy regimeout to 2 years of followup. After 2 years, there was an apparent adverseeffect (increased mortality) of performing an exercise treatment regimein heart failure patients with elevated soluble ST2 levels compared tothe usual care (not performing an exercise therapy regime) patients.

The ST2 analysis cutpoint of 35 ng/mL was selected by choosing a PresageST2 Assay concentration value above the 90th and below the 95thpercentile of the group. These reference values were subsequentlyconfirmed to be consistent with an additional 3,450 subjects measured ina large observational population study, the Framingham Offspring Cohort(Wang et al. 2004) by showing that ST2 values are 32.9 ng/ml and 37.3ng/ml at the 90th and 95th percentiles respectively, bracketing andconfirming the selected 35 ng/ml value.

The relationship between the mortality risk of performing an exercisetherapy regime and soluble ST2 levels in heart failure patients is alsoillustrated in FIG. 4. The data in FIG. 4 show the calculated hazardratio for mortality within 1 year in heart failure subjects performingan exercise treatment regime to heart failure subjects not performing anexercise treatment regime having different soluble ST2 levels. Thesedata show that, as soluble ST2 levels increase there is a steadyincrease in the hazard ratio, reflecting worse survival, and that alevel of soluble ST2 between 55 and 60 ng/mL has a hazard ratio of 1.0.These data indicated that heart failure patients with soluble ST2 levelsbelow 55-60 ng/mL are likely to benefit from performing an exercisetherapy regime, with greater benefit achieved at lower soluble ST2levels. Heart failure patients with soluble ST2 levels above 55-60 ng/mLare not likely to achieve a therapeutic benefit from performing anexercise therapy regime and are at increased risk of experiencing anadverse outcome (e.g., mortality).

OTHER EMBODIMENTS

It is to be understood that while the invention has been described inconjunction with the detailed description thereof, the foregoingdescription is intended to illustrate and not limit the scope of theinvention, which is defined by the scope of the appended claims. Otheraspects, advantages, and modifications are within the scope of thefollowing claims.

What is claimed is:
 1. A method of treating a subject having heartfailure, the method comprising: (a) performing an immunoassay todetermine a level of soluble ST2 in a biological sample from a subjecthaving heart failure at a first time point before the start of exercisetherapy; (b) beginning exercise therapy in a subject having a decreasedlevel of soluble ST2 in the biological sample at the first time point ascompared to an efficacy reference level; (c) performing an immunoassayto determine a level of soluble ST2 in a biological sample from thesubject undergoing exercise therapy at a second time point after thestart of exercise therapy and after the first time point; (d)identifying a subject having a decreased level of soluble ST2 in thebiological sample at the second time point as compared to a riskreference level; and (e) continuing to treat the identified subject withexercise therapy.
 2. The method of claim 1, wherein the biologicalsamples at the first and second time points comprise blood, plasma, orserum.
 3. The method of claim 1, wherein the subject ishypercholesterolemic, hypertriglyceridemic, hyperlidemic, a smoker,hypertensive, or has a body mass index of greater than
 30. 4. The methodof claim 1, further comprising determining a level of cardiac troponin,B type natriuretic peptide, atrial natriuretic peptide, or C-reactiveprotein in the biological sample at the first time point or in thebiological sample at the second time point.
 5. The method of claim 1,wherein the subject is previously diagnosed as having heart failure. 6.The method of claim 1, wherein the immunoassay is an enzyme-linkedimmunosorbent assay (ELISA).
 7. The method of claim 1, wherein one orboth of the efficacy reference level and the risk reference level is athreshold level of soluble ST2.
 8. The method of claim 1, wherein theimmunoassay in step (a) and the immunoassay in step (c) are performedusing one or both of (i) an antibody produced by the hybridoma depositedat the American Type Culture Collection (ATCC) and designated by thePatent Deposit Designation PTA-10431, and (ii) an antibody produced bythe hybridoma deposited at the ATCC and designated by the Patent DepositDesignation PTA-10432.
 9. A method of treating a subject having heartfailure, the method comprising: (a) performing an immunoassay todetermine a level of soluble ST2 in a biological sample from a subjecthaving heart failure at a first time point before the start of exercisetherapy; (b) beginning exercise therapy in a subject having a decreasedlevel of soluble ST2 in the biological sample at the first time point ascompared to an efficacy reference level; (c) performing an immunoassayto determine a level of soluble ST2 in a biological sample from thesubject undergoing exercise therapy at a second time point after thestart of exercise therapy and after the first time point; (d)identifying a subject having an elevated level of soluble ST2 in thebiological sample at the second time point as compared to the riskreference level; and (e) treating the identified subject with atreatment of heart failure that does not include exercise therapy,wherein the treatment comprises a therapeutically effective amount of arenin-angiotensin-aldosterone system inhibitor and a therapeuticallyeffective amount of a beta-adrenergic receptor blocker.
 10. The methodof claim 9, wherein the biological samples at the first and second timepoints comprise blood, plasma, or serum.
 11. The method of claim 9,wherein the subject is hypercholesterolemic, hypertriglyceridemic,hyperlidemic, a smoker, hypertensive, or has a body mass index ofgreater than
 30. 12. The method of claim 9, further comprisingdetermining a level of cardiac troponin, B type natriuretic peptide,atrial natriuretic peptide, or C-reactive protein in the biologicalsample at the first time point or in the biological sample at the secondtime point.
 13. The method of claim 9, wherein the subject is previouslydiagnosed as having heart failure.
 14. The method of claim 9, whereinthe immunoassay is an enzyme-linked immunosorbent assay (ELISA).
 15. Themethod of claim 9, wherein one or both of the efficacy reference leveland the risk reference level is a threshold level of soluble ST2.
 16. Amethod of treating a subject having heart failure, the methodcomprising: (a) performing an immunoassay to determine a level ofsoluble ST2 in a biological sample from a subject having heart failureat a first time point after the start of exercise therapy; (b)continuing exercise therapy in a subject having a decreased level ofsoluble ST2 in the biological sample at the first time point as comparedto an efficacy reference level; (c) performing an immunoassay todetermine a level of soluble ST2 in a biological sample from the subjectundergoing exercise therapy at a second time point after the start ofexercise therapy and after the first time point; (d) identifying asubject having a decreased level of soluble ST2 in the biological sampleat the second time point as compared to a risk reference level; and (e)continuing to treat the identified subject with exercise therapy. 17.The method of claim 16, wherein the biological samples at the first andsecond time points comprise blood, plasma, or serum.
 18. The method ofclaim 16, wherein the subject is hypercholesterolemic,hypertriglyceridemic, hyperlidemic, a smoker, hypertensive, or has abody mass index of greater than
 30. 19. The method of claim 16, furthercomprising determining a level of cardiac troponin, B type natriureticpeptide, atrial natriuretic peptide, or C-reactive protein in thebiological sample at the first time point or in the biological sample atthe second time point.
 20. The method of claim 16, wherein the subjectis previously diagnosed as having heart failure.
 21. The method of claim16, wherein the immunoassay is an enzyme-linked immunosorbent assay(ELISA).
 22. The method of claim 16, wherein one or both of the efficacyreference level and the risk reference level is a threshold level ofsoluble ST2.
 23. The method of claim 16, wherein the immunoassay in step(a) and the immunoassay in step (c) are performed using one or both of(i) an antibody produced by the hybridoma deposited at the American TypeCulture Collection (ATCC) and designated by the Patent DepositDesignation PTA-10431, and (ii) an antibody produced by the hybridomadeposited at the ATCC and designated by the Patent Deposit DesignationPTA-10432.
 24. A method of treating a subject having heart failure, themethod comprising: (a) performing an immunoassay to determine a level ofsoluble ST2 in a biological sample from a subject having heart failureat a first time point after the start of exercise therapy; (b)continuing exercise therapy in a subject having a decreased level ofsoluble ST2 in the biological sample at the first time point as comparedto an efficacy reference level; (c) performing an immunoassay todetermine a level of soluble ST2 in a biological sample from the subjectundergoing exercise therapy at a second time point after the start ofexercise therapy and after the first time point; and (d) identifying asubject having an elevated level of soluble ST2 in the biological sampleat the second time point as compared to the risk reference level; and(e) treating the identified subject with a treatment of heart failurethat does not include exercise therapy, wherein the treatment comprisesa therapeutically effective amount of renin-angiotensin-aldosteronesystem inhibitor and a therapeutically effective amount of abeta-adrenergic receptor blocker.
 25. The method of claim 24, whereinthe biological samples at the first and second time points compriseblood, plasma, or serum.
 26. The method of claim 24, wherein the subjectis hypercholesterolemic, hypertriglyceridemic, hyperlidemic, a smoker,hypertensive, or has a body mass index of greater than
 30. 27. Themethod of claim 24, further comprising determining a level of cardiactroponin, B type natriuretic peptide, atrial natriuretic peptide, orC-reactive protein in the biological sample at the first time point orin the biological sample at the second time point.
 28. The method ofclaim 24, wherein the subject is previously diagnosed as having heartfailure.
 29. The method of claim 24, wherein the immunoassay is anenzyme-linked immunosorbent assay (ELISA).
 30. The method of claim 24,wherein one or both of the efficacy reference level and the riskreference level is a threshold level of soluble ST2.